Neural circuits expressing the serotonin 2C receptor regulate memory in mice and humans

Declined memory is a hallmark of Alzheimer’s disease (AD). Experiments in rodents and human postmortem studies suggest that serotonin (5-hydroxytryptamine, 5-HT) plays a role in memory, but the underlying mechanisms are unknown. Here, we investigate the role of 5-HT 2C receptor (5-HT2CR) in regulating memory. Transgenic mice expressing a humanized HTR2C mutation exhibit impaired plasticity of hippocampal ventral CA1 (vCA1) neurons and reduced memory. Further, 5-HT neurons project to and synapse onto vCA1 neurons. Disruption of 5-HT synthesis in vCA1-projecting neurons or deletion of 5-HT2CRs in the vCA1 impairs neural plasticity and memory. We show that a selective 5-HT2CR agonist, lorcaserin, improves synaptic plasticity and memory in an AD mouse model. Cumulatively, we demonstrate that hippocampal 5-HT2CR signaling regulates memory, which may inform the use of 5-HT2CR agonists in the treatment of dementia.


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Figs. S1 to S8 Tables S1 and S2 Supplementary Figures Fig. S1.No off-target mutation in Htr2c F327L mice.Genomic sequence containing the potential off-target sites identified by Cas-OFFinder were PCR-amplified and sequenced.The grey highlight in each representative sequence trace is the region with potential off-target, but no mutation was identified in all these regions.Related to Fig. 1.

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Fig. S2.5-HT2CR regulates memory.(A) Body weight of male WT and Htr2c F327L/Y mice (6 months) at the time of memory tests.Results shown as mean ± SEM with individual data points (N=14 or 19 mice).(B) Latency to reach the platform during the 12 RAWM training trials.Results shown as mean ± SEM. (C) Swim speed during the RAWM.Results shown as mean ± SEM with individual data points.(D) Performance in the fear conditioning test reported as freezing time in response to contextual cues.Results shown as mean ± SEM. (E) Latency to lick hind paw or jump of male WT and Htr2c F327L/Y mice (6 months) in the hot plate test.Results shown as mean ± SEM with individual data points (N=13 or 15 mice).(F) Body weight of female WT and Htr2c F327L/+ mice (6 months) at the time of memory tests.Results shown as mean ± SEM with individual data points (N=8 or 11 mice).(G) Latency to reach the platform

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Fig. S3.5-HT2CR regulates synaptic plasticity in vCA1 neurons.(A) Left panel: Representative current clamp traces in vCA1 neurons from WT mice treated with vehicle (5 seconds puff).Right panel: Percentage of neurons activated by vehicle puff or were irresponsive.(N= 9 neurons from 3 mice).(B) Left panel: Representative current clamp traces in dCA1 neurons from WT mice treated with lorcaserin (100 µM, 5 seconds puff).Right panel: Percentage of neurons activated by lorcaserin puff or were irresponsive.(N= 12 neurons from 3 mice).(C) Representative EPSC traces before and after LTP induction (with 4 HFS) in vCA1 neurons from WT or Htr2c F327L/Y mice.(D) Magnitude of EPSC elevations before and after LTP induction.(E) Averaged EPSC elevations in the first minute after 3 rd pair of highfrequency stimulation in (D).Results shown as mean ± SEM with individual data points (N=

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Fig. S4.5-HT2CR in the vCA1 regulates memory.(A) Body weight of male control and Htr2c vCA1-KO mice (6 months) at the time of memory tests.Results shown as mean ± SEM with individual data points (N=9 or 10 mice).(B) Latency to reach the platform during the 12 RAWM training trials.Results shown as mean ± SEM. *, P<0.05 in two-way ANOVA analysis.(C) Swim speed during the RAWM.Results shown as mean ± SEM with individual data points.(D) Performance in the fear conditioning test reported as freezing time in response to contextual cues.Results shown as mean ± SEM. (E) A schematic illustration of the open field test.(F) Number of entries to the center of the open field.Results shown as mean ± SEM. (G) Time spent in the center of the open field.Results shown as mean ± SEM. (H) Time spent in the margin of the open field.Results shown as mean ± SEM.Related to Fig. 2.

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Fig. S6.Midbrain 5-HT neurons project to and activate vCA1 neurons.(A) Percentage of tested vCA1 neurons showing light-evoked EPSCs.(B) Representative EPSC traces induced by blue light stimulation (473 nM, 40 mW, 10 ms pulse) in the absence or the presence of various inhibitors.(C-D) Amplitude (C) and latency (D) of light-evoked EPSC.Results are shown as mean ± SEM with individual data points.***, P<0.001 in one way ANOVA followed by Tukey's test (N= 10 neurons from 3 mice per group).(E) Percentage of tested vCA1 neurons showing light-evoked IPSCs.(F) Representative IPSC traces induced by blue light stimulation in the absence or the presence of glutamate receptor inhibitors (30 μM CNQX and 30 μM D-AP5) or the GABAA receptor inhibitor (50 μM bicuculline).(G-H) Amplitude (G) and latency (H) of light-evoked IPSC.Results are shown as mean ± SEM with individual data points.***, P<0.001 in one way ANOVA followed by Tukey's test (N= 5 neurons from 3 mice per group).Related to Fig. 3.

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Fig. S7.5-HTergic projections to the vCA1 regulate memory.(A) Body weight of male control and Di retro-vCA1-Tph2 mice (4 months) at the time of memory tests.Results shown as mean ± SEM with individual data points (N=8 mice).(B) Latency to reach the platform during the 12 RAWM training trials.Results shown as mean ± SEM. (C) Swim speed during the RAWM.Results shown as mean ± SEM with individual data points.(D) Performance in the fear conditioning test reported as freezing time in response to contextual cues.Results shown as mean ± SEM. (E) Representative microscopic images showing Tph2-expressing neurons in the DRN, MRN and CRN of control and Tph2 retro-vCA1-KO mice; scale bar = 100 µm.(F-H) Quantification of neurons that were Tph2 positive in the DRN (F), the MRN (G) and the CRN (H).Results are shown as mean ± SEM with individual points.*, P<0.05 and **, P<0.01 in two-tailed unpaired test (N= 3 mice per group).(I) Body weight of male control and Tph2 retro- vCA1-KO mice (6 months) at the time of memory tests.Results shown as mean ± SEM with individual data points (N=10 or 14 mice).(J) Latency to reach the platform during the 12 RAWM training trials.Results shown as mean ± SEM. (K) Swim speed during the RAWM.Results shown as mean ± SEM with individual data points.(L) Performance in the fear conditioning test reported as freezing time in response to contextual cues.Results shown as mean ± SEM.Related to Fig. 4.